The immune system is characterized by the ability to respond to infectious agents without mounting a destructive response against self-tissues. The first line of defense to invading pathogens is represented by the innate immune response that detects and limits the infection. From the innate response, it originates the adaptive immune response, which discriminates among pathogens and gives rise to memory.
Dendritic cells (DC) are extremely versatile professional antigen presenting cells (APCs) involved in the initiation of both innate and adaptive immunity and in the differentiation of regulatory T cells required for the maintenance of self-tolerance. Key functions of DC include uptake and processing of antigens and priming of naive T cells1, functions which are segregated in time. Immature resting DC located in non-lymphoid tissues, such as skin and mucosae, take up antigen. Immature DC are highly fagocitic and continuously internalize soluble and particulate antigens that are processed and presented to T cells. The interaction of immature DC with T cells induces an abortive T cell activation with the induction of T cell anergy or the differentiation of regulatory T cells.
Mature DC loaded with antigens and capable of priming T cells migrate from non-lymphoid tissues to the T cell area of lymph nodes or spleen. Thus, when immature DC come in contact with inflammatory stimuli, they undergo a maturation process that transforms them from phagocytic and migratory cells to non-phagocytic, highly efficient stimulators of naive T cell responses. Mature DC are programmed to undergo apoptotic death in 9-10 days.
Inflammatory and microbial stimuli induce the DC maturation process that concludes 24 hours later. Mature DC express at the cell surface high levels of stable peptide-MHC complexes and costimulatory molecules and are capable of prime naive T cells. During the process of differentiation, DC undergo intermediate maturational stages in which they express, with a strictly defined kinetic, cytokines and cell surface molecules critical for activation and control of, first, innate and, then, adaptive immune responses.
Immature monocyte derived human DC (hMDC) or immature bone marrow-derived mouse DC (mBMDC) can be induced to mature in vitro using many different stimuli, including inflammatory cytokines, bacteria cell products like lipopolysaccharide (LPS) and lipoteichoic acid (LTA), bacterial DNA and double-stranded viral RNA, and live bacteria. This last stimulus represents one of the most potent catalysts of DC terminal differentiation process in the mouse, inducing a rapid and effective DC phenotypic and functional maturation4.
The extent and the type of innate and adaptive responses DC activate depend on the type of stimulus they receive. DC are, actually, able to distinguish different pathogens since they express pattern-recognition receptors (PRRs) that interact with specific microorganism molecular structures called pathogens-associated molecular patterns (PAMPs). These constitutive and conserved microbial structures are absent in host mammalian cells and represent the signature of different microorganisms.
Well-defined PRRs are Toll like receptors (TLRs). The stimulation of different TLRs at the DC surface results in the activation of different signaling pathways and in the induction of different maturation processes that influence the outcome of adaptive immunity. In this sense DC are able to respond in a pathogen-specific way.
The transcription profile, transcriptome, is a major determinant of cellular phenotype and function. Differences in gene expression are indicative of morphological, phenotypical and functional changes induced in a cell by environmental factors and perturbations. Microarrays have been successfully applied to identify genes that discriminate between Th1 and Th2 functions in humans and anergic and activated B cells in mice. Micro-array technology is, thus, a valid approach to investigate possible differences induced in particular cell types by diverse external factors.
Concerning DC, a transcriptional profile of immature and LPS-matured human monocyte-derived DC has been carried out revealing 225 differentially expressed genes in the two situations out of a total of 10,962 genes screened. These genes mainly consisted of chemokines (RANTES, ELC, PARK, MDC and TARC) and chemokine receptors (CCR7), enzymes (such as germinal center kinase-related protein kinase), and IFN-inducible proteins (lipase A, CD52, CD11b, CD23, and glucose 6 phosphatase). Nevertheless a comparison between different stimuli in their efficiency in inducing DC maturation has never been performed.
Immunosuppression represents a common outcome of viral infections. The down-regulation of immune responses imparts the infecting pathogens the opportunity to escape immune surveillance and thus maximizes their chances to survive within their host, to replicate and be transmitted as required. The generalized immunosuppression caused by viral infection is often associated with secondary infections with unrelated viral and/or bacterial pathogens and, as such, represents a serious clinical problem. Immunosuppression may also accompany the onset of tumors, again as a means for tumors to evade immune responses and reduce the chances of being eliminated. Understanding the mechanisms involved in the induction of immunosuppression is therefore a crucial step towards the development of better immunotherapies.
Cytomegalovirus (CMV) is one of three viral pathogens of humans known to induce transient, but profound immunosuppression. Unlike measles and HIV, the mechanisms underlying CMV-induced immunosuppression remain poorly understood despite the availability of a unique animal model. Murine cytomegalovirus (MCMV) infection is widely utilized as a model for human CMV infection. Human CMV (HCMV) causes serious complications in immunocompromised hosts such as newborns, transplant patients and AIDS patients. Since CMV infection is species specific, there are no experimental animal models to study HCMV pathogenesis. However, because of the similarity in structure and biology between HCMV and MCMV, the latter provides a unique model of human disease and importantly it permits the study of in vivo infection of the natural host.
In humans, CMV infection causes morbidity and mortality amongst neonates and immunosuppressed individuals. In immunocompetent hosts however, CMV can establish a persistent infection without causing overt disease. The ability to persist and establish a stable relationship with its host is critical to the survival of CMV and strongly underlies its success at evading host defense mechanisms. Amongst the strategies used by CMV to subvert normal defense mechanisms is the “hijacking” of cellular gene products that play critical roles in antiviral responses.
MCMV is capable of subverting the immune system by multiple mechanisms, including down regulation of MHC-I and II molecules, synthesis of chemokine homologues and production of a viral homologue of cellular MHC-I. Studies with mutant viruses lacking specific viral ORFs have clearly demonstrated the effects of MCMV proteins in the regulation of T cell activity and the inhibition of NK cell responses. Studies in murine models have also helped defining the role of specific cellular subsets, with monocytes and macrophages having been shown to be important in viral dissemination and pathogenicity. Interestingly, recent studies have shown that HCMV infects monocyte derived dendritic cells (DC). Importantly, however, there are no reports on the role of DC in MCMV infection, this being the only system that allows the analysis of the biological significance of DC infection in vivo.
There is a need in the art to develop methods and compositions useful for regulating DC, especially in association with DC related immune responses.